Hypoxia regulated conditionally silenced aav expressing angiogenic inducers

Abstract

Methods and compositions for the treatment of hypoxia associated disorders by directional angiogenesis/arteriogenesis. Conditionally silenced vectors expressing a therapeutic molecule under hypoxic conditions avoid chaotic vascularization and allow for the orderly growth of new vessels into damaged tissue.

1 - 34 . (canceled) 
     
     
         35 . An adeno-associated virus (AAV) particle comprising at least one hypoxia response element (HRE), at least one neuron restrictive silencer element (NRSE) that binds neuron restrictive silencer factor, a gene encoding vascular endothelial growth factor (VEGF), and a phosphoglycerate kinase promoter upstream of the gene and in operable linkage with the gene, the at least one HRE and the at least one NRSE. 
     
     
         36 . The AAV particle according to  claim 35 , wherein the AAV particle comprises a combined cassette of HRE and NRSE. 
     
     
         37 . The AAV particle according to  claim 35 , wherein the AAV particle further comprises at least one FROG element and/or at least one TOAD element. 
     
     
         38 . The AAV particle according to  claim 37 , wherein the AAV particle comprises a combined cassette of HRE, NRSE, FROG and/or TOAD. 
     
     
         39 . The AAV particle according to  claim 37 , wherein the at least one FROG element consists of a nucleotide sequence selected from the group consisting of: SEQ ID NOS: 3 and 4, and the at least one TOAD element consists of a nucleotide sequence selected from the group consisting of: SEQ ID NOS: 1 and 2. 
     
     
         40 . The AAV particle according to  claim 37 , wherein the at least one FROG element and the at least one TOAD element are a nucleotide sequence selected from the group consisting of: SEQ ID NOS: 5 and 6. 
     
     
         41 . The AAV particle according to  claim 35 , wherein the tissue is cardiac tissue or limb tissue. 
     
     
         42 . A method of treating a patient having critical limb ischemia comprising administering to the patient a composition comprising an adeno-associated virus (AAV) particle comprising at least one hypoxia response element (HRE) and at least one neuron restrictive silencer element (NRSE) that binds neuron restrictive silencer factor, a gene encoding vascular endothelial growth factor (VEGF), and a phosphoglycerate kinase promoter upstream of the gene and in operable linkage with the gene, the at least one HRE and the at least one NRSE, in a therapeutically effective controlled amount for inhibiting necrosis and increasing blood flow in the ischemic or hypoxic limb tissue. 
     
     
         43 . The method according to  claim 42 , wherein the AAV particle comprises a combined cassette of HRE and NRSE. 
     
     
         44 . The method according to  claim 42 , wherein the AAV particle further comprises at least one FROG element and/or at least one TOAD element. 
     
     
         45 . method according to  claim 44 , wherein the AAV particle comprises a combined cassette of HRE, NRSE, FROG and/or TOAD. 
     
     
         46 . The method according to  claim 44 , wherein the at least one FROG element consists of a nucleotide sequence selected from the group consisting of: SEQ ID NOS: 3 and 4, and the at least one TOAD element consists of a nucleotide sequence selected from the group consisting of: SEQ ID NOS: 1 and 2. 
     
     
         47 . The method according to  claim 44 , wherein the at least one FROG element and the at least one TOAD element are a nucleotide sequence selected from the group consisting of: SEQ ID NOS: 5 and 6. 
     
     
         48 . An adeno-associated virus (AAV) particle comprising at least one NF-κB inflammatory response element (IRE), at least one neuron restrictive silencer element (NRSE) that binds neuron restrictive silencer factor, a gene encoding vascular endothelial growth factor (VEGF), and a phosphoglycerate kinase promoter upstream of the gene and in operable linkage with the gene, the at least one IRE and the at least one NRSE. 
     
     
         49 . The AAV particle according to  claim 48 , wherein the AAV particle further comprises at least one FROG element and/or at least one TOAD element. 
     
     
         50 . A method of treating a patient having critical limb ischemia comprising administering to the patient a composition comprising an adeno-associated virus (AAV) particle comprising at least one NF-κB inflammatory response element (IRE) and at least one neuron restrictive silencer element (NRSE) that binds neuron restrictive silencer factor, a gene encoding vascular endothelial growth factor (VEGF), and a phosphoglycerate kinase promoter upstream of the gene and in operable linkage with the gene, the at least one IRE and the at least one NRSE, in a therapeutically effective controlled amount for inhibiting necrosis and increasing blood flow in the ischemic or hypoxic limb tissue. 
     
     
         51 . The method according to  claim 50 , wherein the AAV particle further comprises at least one FROG element and/or at least one TOAD element.